Carcinoembryonic antigen (CEA) is a 180,000-dalton glycoprotein expressed on most gastrointestinal carcinomas and many other human carcinoma types. A 2.4-kb cDNA clone, containing the complete coding sequence, was isolated from a human colon tumor cell library and inserted into a vaccinia virus genome. This newly developed construct was characterized by Southern blotting, DNA hybridization studies and polymerase chain reaction analysis. The CEA gene was stably integrated in the vaccinia virus thymidine kinase gene. The recombinant was efficiently replicated upon serial passages in cell cultures and in animals. The recombinant virus expresses, on the surface of infected cells, a protein product recognized by a monoclonal antibody (COL-1) directed against CEA. Immunization of mice with the vaccinia construct was found to elicit a humoral immune response against CEA. Pilot studies also demonstrated that the administration of the recombinant CEA vaccinia construct was able to greatly reduce the growth in mice of a syngenic murine colon adenocarcinoma which had been transduced with the human CEA gene. The use of this new recombinant CEA vaccinia construct may thus provide an approach in the specific active immunotherapy of human gastrointestinal cancer and other CEA expressing carcinoma types.